Analysis of Neuroactive Amines in Fermented Beverages Using a Portable Microchip Capillary Electrophoresis System Christine N. Jayarajah, Alison M. Skelley, † Angela D. Fortner, and Richard A. Mathies* Department of Chemistry, University of California, Berkeley, California 94720 A portable microfabricated capillary electrophoresis (CE) instrument is used for the determination of neurologically active biogenic amines, especially tyramine and hista- mine, in fermented beverages. The target molecules are labeled on their primary amino groups with fluorescamine in a 10-min reaction, and the samples analyzed directly, producing a detailed electropherogram in only 120 s on a microfabricated glass CE device containing 21.4-cm- long separation channels. Tyramine was found mainly in red wines at <1-3.4 mg/L, while the histamine content of these samples ranged from 1.8 to 19 mg/L. The highest levels of histamine (20-40 mg/L) were found in sake. The analysis of samples drawn from grape crush through malolactic fermentation in four varieties of zinfandel red wines revealed that histamine and tyramine are produced during yeast and malolactic fermentation, respectively. Following malolactic fermentation, the histamine content in these samples ranged from 3.3 to 30 mg/L, and the tyramine content ranged from 1.0 to 3.0 mg/L. This highly sensitive and rapid lab-on-a-chip analysis method estab- lishes the feasibility of monitoring neurologically active amine content and potentially other chemically and aller- genically important molecules in our food supply. Tyramine and histamine, produced by the decarboxylation of tyrosine and histidine, are among the most harmful of the biogenic amines found in fermented beverages (Figure 1). These amines, produced as degradation products resulting from microbial activ- ity, are found widely in fermented foods and beverages, meat, fish, and diary products. 1-7 Biogenic amines such as histamine, tyramine, and phenylethylamine are known to induce nausea, headaches, and respiratory disorders in sensitive individuals, particularly when accompanied by alcohol and acetaldehyde. 8 These amines are normally metabolized by amine oxidases to keep their steady-state concentrations low. However, for individuals with reduced monoamine oxidase (MAO) activity or expression and for individuals taking MAO inhibitors, ingestion of foods contain- ing large amounts of tyramine can lead to transient hypertension, hypertensive crisis, and panic attacks. 9 This response occurs because tyramine that is not deaminated is converted to octo- pamine when taken up in sympathetic nerve terminals where it displaces norepinephrine (NE) from storage vesicles. A portion of this NE diffuses out of the nerve to react with receptors causing hypertension and other sympathomimetic effects. 10 Analogously, histamine in wine can induce headaches in patients suffering from reduced or lack of diamine oxidase activity. The treatment of choice for patients with histamine or tyramine intolerance and chronic headache is a histamine- and tyramine-free diet. 11 For these reasons, the development of a fast, accurate, point-of- consumption (POC) method to measure biogenic amine concen- trations in foods would be valuable. Previous methods to determine the biogenic amine content in foods involve conventional chromatographic separations with extensive and complex derivatization protocols and sample processing. High-performance liquid chromatography (HPLC) methods typically include pre- or postcolumn derivatization and fluorometric detection of the corresponding derivatives. 12 The commonly used derivatization reagent is o-phthaldialdehyde in the presence of 2-mercaptoethanol. 13,14 A recent method for the simultaneous HPLC analysis of biogenic amines, amino acids, and ammonium ion in wine and beer samples as aminoenone deriva- tives involves reaction with the derivatization reagent diethyl ethoxymethylenemalonate in methanolic alkaline medium. 15 Millan et al. have developed a more rapid liquid chromatographic- * To whom correspondence should be addressed. Phone: (510) 642-4192. Fax: (510) 642-3599. E-mail: rich@zinc.cchem.berkeley.edu. † Present address: Research Laboratory of Electronics, Massachusetts Institute of Technology, Cambridge, MA 02139. (1) McCabe-Sellers, B. J.; Staggs, C. G.; Bogle, M. L. J. Food Comp. Anal. 2006, 19, S58-S65. (2) Margalit, Y. Concepts in Wine Chemistry; The Wine Appreciation Guild: San Francisco, 2004. (3) Moret, S.; Smela, D.; Populin, T.; Conte, L. S. Food Chem. 2005, 89, 355- 361. (4) Ruiz-Capillas, C.; Jimenez-Colmenero, F. Crit. Rev. Food Sci. Nutr. 2004, 44, 489-499. (5) Kalac, P.; Krizek, M. J. Inst. Brew. 2003, 109, 123-128. (6) Jansen, S. C.; van Dusseldorp, M.; Bottema, K. C.; Dubois, A. E. J. Ann. Allergy Asthma Immunol. 2003, 91, 233-241. (7) Lange, J.; Wittmann, C. Anal. Bioanal. Chem. 2002, 372, 276-283. (8) Silla Santos, M. H. Int. J. Food Microbiol. 1996, 29, 213-231. (9) Caston, J. C.; Eaton, C. L.; Gheorghiu, B. P.; Ware, L. L. South Carolina Med. J. 2002, 98, 187-192. (10) Mayer, S. E. In The Pharmacological Basis of Therapeutics, 6th ed.; Gilman, A. G., Goodman, L. S., Gilman, A., Eds.; Macmillan Publishing Co., Inc.: New York, 1980; pp 74-75. (11) McCabe, B. J. In Handbook of Food and Drug Interactions; McCabe, B. J., Frankel, E. H., Wolfe, J. J., Eds.; CRC Press: Boca Raton, FL, 2003; pp 457- 459. (12) Molnar-Perl, I. J. Chromatogr., A 2003, 987, 291-309. (13) Hanczko, R.; Molnar-Perl, I. Chromatographia 2003, 57, S103-S113. (14) Kutlan, D.; Molnar-Perl, I. J. Chromatogr., A 2003, 987, 311-322. (15) Gomez-Alonso, S.; Hermosin-Gutierrez, I.; Garcia-Romero, E. J. Agric. Food Chem. 2007, 55, 608-613. Anal. Chem. 2007, 79, 8162-8169 8162 Analytical Chemistry, Vol. 79, No. 21, November 1, 2007 10.1021/ac071306s CCC: $37.00 © 2007 American Chemical Society Published on Web 09/25/2007